Mycobacterium tuberculosis (Mtb) remains one of the world's most prevalent and deadly bacterial pathogens, as over 2 billion humans are currently infected and 3 million die annually. The widely used M. bovis Bacille-Calmette-Gurin (BCG) vaccine offers variable protection against Mtb infection in adults, making the development of improved vaccines that generate strong cellular immune responses a priority. CD1 molecules are a third lineage of antigen presenting molecules which are specialized in presenting lipid antigens to T cells. In humans, this family consists of group 1 CD1 molecules CD1a, -b, and -c, and the group 2 molecule CD1d. Mice lack group 1 CD1, but express group 2 CD1. While murine CD1d has been extensively studied, the role group 1 CD1-restricted T cells play in host immunity is poorly understood due to the lack of a suitable small animal model. To study the in vivo function of group 1 CD1-restricted T cells, our group has developed a novel animal model by introducing the entire human group 1 CD1 locus into the mouse. Human group 1 CD1 transgenic (hCD1Tg) mice express CD1a, CD1b, and CD1c in a pattern similar to that seen in humans. In this project, we will investigate the in vivo function of group 1 CD1-restricted T cells during Mtb infections. There is a lack of evidence about the relative contribution of individual Mtb lipid-specific group 1 CD1- restricted T cells to protective immunity against Mtb. To bridge this gap, in Specific Aim 1, we will screen known Mtb lipid antigens to identify immunodominant lipid antigens during Mtb infection. We will use the immunodominant lipid antigens in an immunization protocol to evaluate the protective capacity of lipid-specific group 1 CD1-restricted T cells. The in vivo dynamics of Mtb-specific group 1 CD1 T cells during infection is not well understood. In Specific Aim 2, we will isolate APCs from different tissues and measure group 1 CD1 surface expression during Mtb infection. Furthermore, we will use a tetramer-based approach to evaluate the tissue location, kinetics, and effector function of lipid-specific group 1 CD1-restricted T cells during Mtb infection. Our results will provide important information about where lipid-specific group 1 CD1-restricted T cells are activated in vivo and their phenotype. Lastly, in Specific Aim 3, we will evaluate the role of autoreactive group 1 CD1-restricted T cells in immune defense or immunopathology of Mtb. Recent data suggests a large proportion of group 1 CD1- restricted T cells are autoreactive. Moreover, we observed, in preliminary experiments, a large population of group 1 CD1-restricted T cells persist in the lung after the lipid-specific group 1-restricted T cell response diminished. We will evaluate the tissue distribution, surface phenotype and functional properties of these cells to gain an understanding of their role in immunopathology.